HGNC Approved Gene Symbol:RRAGA
Cytogenetic location:9p22.1 Genomic coordinates(GRCh38) :9:19,049,427-19,051,025 (from NCBI)
Using mouse Raga to screen a fetal brain cDNA library, followed by RACE,Schurmann et al. (1995) cloned human RAGA. The deduced 313-amino acid protein shares similarity with RAS (HRAS;190020) and contains phosphate/magnesium-binding motifs PM1, PM2, and PM3, and the guanine nucleotide-binding motif G1. An additional guanine nucleotide-binding motif, G2, is strikingly different from those found in all other RAS homologs. The C-terminal domain contains a putative transmembrane region. Northern blot analysis of rat tissues detected highest expression in adrenal gland.
Using adenovirus E3-14.7K protein in a yeast 2-hybrid screen of a HeLa cell cDNA library,Li et al. (1997) cloned RRAGA, which they called FIP1. They noted that the protein contains 2 putative myristoylation sites, several putative phosphorylation sites, and 2 domains homologous to bacterial metalloproteases. In transfected mouse fibroblasts, FIP1 colocalized with adenovirus E3-4.7K in the cytoplasm, especially near the nuclear membrane and in discrete foci on or near the plasma membrane; FIP1 also localized without E3-4.7K in the nucleus. Northern blot analysis detected FIP1 in all human tissues examined, with highest expression in skeletal muscle.
Schurmann et al. (1995) found that recombinant RAGA bound a nonhydrolyzable analog of GTP, but that it lacked intrinsic GTPase activity.
Using a protein pull-down assay with recombinant proteins,Li et al. (1997) confirmed the interaction between FIP1 and adenovirus E3-14.7K. Mutation analysis revealed N- and C-terminal domains of FIP1 that were required for the interaction. In human embryonic kidney cells, TNF-alpha (TNF;191160), a proinflammatory cytokine with antiviral activity, promoted the transient association of FIP1 with phosphoproteins. In mouse fibroblasts, antisense FIP1 RNA inhibited TNF-alpha-induced cytolysis.
The multiprotein mTORC1 protein kinase complex (see601231) is the central component of a pathway that promotes growth in response to insulin, energy levels, and amino acids and is deregulated in common cancers.Sancak et al. (2008) found that the Rag proteins, a family of 4 related small guanosine triphosphatases (GTPases) (RAGA, RAGB (300725), RAGC (608267), and RAGD (608268)) interact with mTORC1 in an amino acid-sensitive manner and are necessary for the activation of the mTORC1 pathway by amino acids. A Rag mutant that was constitutively bound to guanosine triphosphate interacted strongly with mTORC1, and its expression within cells made the mTORC1 pathway resistant to amino acid deprivation. Conversely, expression of a guanosine diphosphate-bound Rag mutant prevented stimulation of mTORC1 by amino acids.Sancak et al. (2008) concluded that the Rag proteins do not directly stimulate the kinase activity of mTORC1, but, like amino acids, promote the intracellular localization of mTOR to a compartment that also contains its activator RHEB (601293).
Bar-Peled et al. (2013) identified the octameric GATOR (GTPase-activating protein (GAP) activity toward RAGs) complex as a critical regulator of the pathway that signals amino acid sufficiency to mTORC1. GATOR is composed of 2 subcomplexes, GATOR1 and GATOR2. Inhibition of the GATOR1 subunits DEPDC5 (614191), NPRL2 (607072), and NPRL3 (600928) makes mTORC1 signaling resistant to amino acid deprivation. In contrast, inhibition of the GATOR2 subunits MIOS (615359), WDR24 (620307), WDR59 (617418), SEH1L (609263), and SEC13 (600152) suppresses mTORC1 signaling, and epistasis analysis shows that GATOR2 negatively regulates DEPDC5. GATOR1 has GAP activity for RAGA and RAGB, and its components are mutated in human cancer. In cancer cells with inactivating mutations in GATOR1, mTORC1 is hyperactive and insensitive to amino acid starvation, and such cells are hypersensitive to rapamycin, an mTORC1 inhibitor. Thus,Bar-Peled et al. (2013) concluded that they had identified a key negative regulator of the RAG GTPases and revealed that, like other mTORC1 regulators, RAG function can be deregulated in cancer.
Using knockdown HEK293 cells and knockout mouse embryonic fibroblasts,Jewell et al. (2015) found that functional RAGA and RAGB were required for activation of lysosomal mTORC1 by leucine. In contrast, ARF1 (103180) was required for activation of lysosomal mTORC1 by glutamine.
Deng et al. (2015) reported that UBC13 (UBE2N;603679) functioned as an E2 ubiquitin-conjugating enzyme and that RNF152 (616512) functioned as an E3 ubiquitin ligase for polyubiquitination of RAGA in HEK293T cells. At least 4 lysines in RAGA were subject to RNF152-dependent attachment of lys63-linked polyubiquitin chains. This polyubiquitination did not result in RAGA degradation, but it promoted interaction of RAGA with GATOR1, which maintains RAGA in the inactive GDP-bound form, thereby inactivating MTORC1.Deng et al. (2015) identified DUB3 (USP17L2;610186) as the predominant deubiquitinating enzyme that removed ubiquitin chains from RAGA.
Cryoelectron Microscopy
Shen et al. (2018) used cryoelectron microscopy to solve structures of GATOR1 and GATOR1-Rag GTPases complexes. GATOR1 adopts an extended architecture with a cavity in the middle; NPRL2 (607072) links DEPDC5 (614191) and NPRL3 (600928), and DEPDC5 contacts the Rag GTPase heterodimer. Biochemical analyses revealed that this GATOR1-Rag GTPases structure is inhibitory, and that at least 2 binding modes must exist between the Rag GTPases and GATOR1. Direct interaction of DEPDC5 with RAGA inhibits GATOR1-mediated stimulation of GTP hydrolysis by RAGA, whereas weaker interactions between the NPRL2-NPRL3 heterodimer and RAGA execute GAP activity.
Hartz (2015) mapped the RRAGA gene to chromosome 9p22.1 based on an alignment of the RRAGA sequence (GenBankX90529) with the genomic sequence (GRCh38).
Efeyan et al. (2013) generated knock-in mice that express a constitutively active form of RagA, RagA(GTP), from its endogenous promoter. RagA(GTP/GTP) homozygous mice developed normally but failed to survive postnatal day 1. When delivered by cesarean section, fasted RagA(GTP/GTP) neonates die almost twice as rapidly as wildtype littermates. Within an hour of birth wildtype neonates strongly inhibit mTORC1 (601231), which coincides with profound hypoglycemia and a decrease in plasma amino acid concentrations. In contrast, mTORC1 inhibition does not occur in RagA(GTP/GTP) neonates, despite identical reductions in blood nutrient amounts. With prolonged fasting, wildtype neonates recover their plasma glucose concentrations, but RagA(GTP/GTP) mice remain hypoglycemic until death, despite using glycogen at a faster rate. The glucose homeostasis defect correlates with the inability of fasted RagA(GTP/GTP) neonates to trigger autophagy and produce amino acids for de novo glucose production. Because profound hypoglycemia does not inhibit mTORC1 in RagA(GTP/GTP) neonates,Efeyan et al. (2013) considered the possibility that the Rag pathway signals glucose as well as amino acid sufficiency to mTORC1. Indeed, mTORC1 is resistant to glucose deprivation in RagA(GTP/GTP) fibroblasts, and glucose, like amino acids, controls its recruitment to the lysosomal surface, the site of mTORC1 activation. Thus, the Rag GTPases signal glucose and amino acid concentrations to mTORC1, and have an unexpectedly key role in neonates in autophagy induction and thus nutrient homeostasis and viability.
Bar-Peled, L., Chantranupong, L., Cherniack, A. D., Chen, W. W., Ottina, K. A., Grabiner, B. C., Spear, E. D., Carter, S. L., Meyerson, M., Sabatini, D. M.A tumor suppressor complex with GAP activity for the Rag GTPases that signal amino acid sufficiency to mTORC1. Science 340: 1100-1106, 2013. [PubMed:23723238,images,related citations] [Full Text]
Deng, L., Jiang, C., Chen, L., Jin, J., Wei, J., Zhao, L., Chen, M., Pan, W., Xu, Y., Chu, H., Wang, X., Ge, X., Li, D., Liao, L., Liu, M., Li, L., Wang, P.The ubiquitination of RagA GTPase by RNF152 negatively regulates mTORC1 activation. Molec. Cell 58: 804-818, 2015. [PubMed:25936802,related citations] [Full Text]
Efeyan, A., Zoncu, R., Chang, S., Gumper, I., Snitkin, H., Wolfson, R. L., Kirak, O., Sabatini, D. D., Sabatini, D. M.Regulation of mTORC1 by the Rag GTPases is necessary for neonatal autophagy and survival. Nature 493: 679-683, 2013. [PubMed:23263183,images,related citations] [Full Text]
Hartz, P. A.Personal Communication. Baltimore, Md. 1/27/2015.
Jewell, J. L., Kim, Y. C., Russell, R. C., Yu, F.-X., Park, H. W., Plouffe, S. W., Tagliabracci, V. S., Guan, K.-L.Differential regulation of mTORC1 by leucine and glutamine. Science 347: 194-198, 2015. [PubMed:25567907,images,related citations] [Full Text]
Li, Y., Kang, J., Horwitz, M. S.Interaction of an adenovirus 14.7-kilodalton protein inhibitor of tumor necrosis factor alpha cytolysis with a new member of the GTPase superfamily of signal transducers. J. Virol. 71: 1576-1582, 1997. [PubMed:8995684,related citations] [Full Text]
Sancak, Y., Peterson, T. R., Shaul, Y. D., Lindquist, R. A., Thoreen, C. C., Bar-Peled, L., Sabatini, D. M.The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science 320: 1496-1501, 2008. [PubMed:18497260,images,related citations] [Full Text]
Schurmann, A., Brauers, A., Massmann, S., Becker, W., Joost, H.-G.Cloning of a novel family of mammalian GTP-binding proteins (RagA, RagB(S), RagB(1)) with remote similarity to the Ras-related GTPases. J. Biol. Chem. 270: 28982-28988, 1995. [PubMed:7499430,related citations] [Full Text]
Shen, K., Huang, R. K., Brignole, E. J., Condon, K. J., Valenstein, M. L., Chantranupong, L., Bomaliyamu, A., Choe, A., Hong, C., Yu, Z., Sabatini, D. M.Architecture of the human GATOR1 and GATOR1-Rag GTPases complexes. Nature 556: 64-69, 2018. [PubMed:29590090,images,related citations] [Full Text]
Alternative titles; symbols
HGNC Approved Gene Symbol: RRAGA
Cytogenetic location: 9p22.1 Genomic coordinates(GRCh38) : 9:19,049,427-19,051,025(from NCBI)
Using mouse Raga to screen a fetal brain cDNA library, followed by RACE, Schurmann et al. (1995) cloned human RAGA. The deduced 313-amino acid protein shares similarity with RAS (HRAS; 190020) and contains phosphate/magnesium-binding motifs PM1, PM2, and PM3, and the guanine nucleotide-binding motif G1. An additional guanine nucleotide-binding motif, G2, is strikingly different from those found in all other RAS homologs. The C-terminal domain contains a putative transmembrane region. Northern blot analysis of rat tissues detected highest expression in adrenal gland.
Using adenovirus E3-14.7K protein in a yeast 2-hybrid screen of a HeLa cell cDNA library, Li et al. (1997) cloned RRAGA, which they called FIP1. They noted that the protein contains 2 putative myristoylation sites, several putative phosphorylation sites, and 2 domains homologous to bacterial metalloproteases. In transfected mouse fibroblasts, FIP1 colocalized with adenovirus E3-4.7K in the cytoplasm, especially near the nuclear membrane and in discrete foci on or near the plasma membrane; FIP1 also localized without E3-4.7K in the nucleus. Northern blot analysis detected FIP1 in all human tissues examined, with highest expression in skeletal muscle.
Schurmann et al. (1995) found that recombinant RAGA bound a nonhydrolyzable analog of GTP, but that it lacked intrinsic GTPase activity.
Using a protein pull-down assay with recombinant proteins, Li et al. (1997) confirmed the interaction between FIP1 and adenovirus E3-14.7K. Mutation analysis revealed N- and C-terminal domains of FIP1 that were required for the interaction. In human embryonic kidney cells, TNF-alpha (TNF; 191160), a proinflammatory cytokine with antiviral activity, promoted the transient association of FIP1 with phosphoproteins. In mouse fibroblasts, antisense FIP1 RNA inhibited TNF-alpha-induced cytolysis.
The multiprotein mTORC1 protein kinase complex (see 601231) is the central component of a pathway that promotes growth in response to insulin, energy levels, and amino acids and is deregulated in common cancers. Sancak et al. (2008) found that the Rag proteins, a family of 4 related small guanosine triphosphatases (GTPases) (RAGA, RAGB (300725), RAGC (608267), and RAGD (608268)) interact with mTORC1 in an amino acid-sensitive manner and are necessary for the activation of the mTORC1 pathway by amino acids. A Rag mutant that was constitutively bound to guanosine triphosphate interacted strongly with mTORC1, and its expression within cells made the mTORC1 pathway resistant to amino acid deprivation. Conversely, expression of a guanosine diphosphate-bound Rag mutant prevented stimulation of mTORC1 by amino acids. Sancak et al. (2008) concluded that the Rag proteins do not directly stimulate the kinase activity of mTORC1, but, like amino acids, promote the intracellular localization of mTOR to a compartment that also contains its activator RHEB (601293).
Bar-Peled et al. (2013) identified the octameric GATOR (GTPase-activating protein (GAP) activity toward RAGs) complex as a critical regulator of the pathway that signals amino acid sufficiency to mTORC1. GATOR is composed of 2 subcomplexes, GATOR1 and GATOR2. Inhibition of the GATOR1 subunits DEPDC5 (614191), NPRL2 (607072), and NPRL3 (600928) makes mTORC1 signaling resistant to amino acid deprivation. In contrast, inhibition of the GATOR2 subunits MIOS (615359), WDR24 (620307), WDR59 (617418), SEH1L (609263), and SEC13 (600152) suppresses mTORC1 signaling, and epistasis analysis shows that GATOR2 negatively regulates DEPDC5. GATOR1 has GAP activity for RAGA and RAGB, and its components are mutated in human cancer. In cancer cells with inactivating mutations in GATOR1, mTORC1 is hyperactive and insensitive to amino acid starvation, and such cells are hypersensitive to rapamycin, an mTORC1 inhibitor. Thus, Bar-Peled et al. (2013) concluded that they had identified a key negative regulator of the RAG GTPases and revealed that, like other mTORC1 regulators, RAG function can be deregulated in cancer.
Using knockdown HEK293 cells and knockout mouse embryonic fibroblasts, Jewell et al. (2015) found that functional RAGA and RAGB were required for activation of lysosomal mTORC1 by leucine. In contrast, ARF1 (103180) was required for activation of lysosomal mTORC1 by glutamine.
Deng et al. (2015) reported that UBC13 (UBE2N; 603679) functioned as an E2 ubiquitin-conjugating enzyme and that RNF152 (616512) functioned as an E3 ubiquitin ligase for polyubiquitination of RAGA in HEK293T cells. At least 4 lysines in RAGA were subject to RNF152-dependent attachment of lys63-linked polyubiquitin chains. This polyubiquitination did not result in RAGA degradation, but it promoted interaction of RAGA with GATOR1, which maintains RAGA in the inactive GDP-bound form, thereby inactivating MTORC1. Deng et al. (2015) identified DUB3 (USP17L2; 610186) as the predominant deubiquitinating enzyme that removed ubiquitin chains from RAGA.
Cryoelectron Microscopy
Shen et al. (2018) used cryoelectron microscopy to solve structures of GATOR1 and GATOR1-Rag GTPases complexes. GATOR1 adopts an extended architecture with a cavity in the middle; NPRL2 (607072) links DEPDC5 (614191) and NPRL3 (600928), and DEPDC5 contacts the Rag GTPase heterodimer. Biochemical analyses revealed that this GATOR1-Rag GTPases structure is inhibitory, and that at least 2 binding modes must exist between the Rag GTPases and GATOR1. Direct interaction of DEPDC5 with RAGA inhibits GATOR1-mediated stimulation of GTP hydrolysis by RAGA, whereas weaker interactions between the NPRL2-NPRL3 heterodimer and RAGA execute GAP activity.
Hartz (2015) mapped the RRAGA gene to chromosome 9p22.1 based on an alignment of the RRAGA sequence (GenBank X90529) with the genomic sequence (GRCh38).
Efeyan et al. (2013) generated knock-in mice that express a constitutively active form of RagA, RagA(GTP), from its endogenous promoter. RagA(GTP/GTP) homozygous mice developed normally but failed to survive postnatal day 1. When delivered by cesarean section, fasted RagA(GTP/GTP) neonates die almost twice as rapidly as wildtype littermates. Within an hour of birth wildtype neonates strongly inhibit mTORC1 (601231), which coincides with profound hypoglycemia and a decrease in plasma amino acid concentrations. In contrast, mTORC1 inhibition does not occur in RagA(GTP/GTP) neonates, despite identical reductions in blood nutrient amounts. With prolonged fasting, wildtype neonates recover their plasma glucose concentrations, but RagA(GTP/GTP) mice remain hypoglycemic until death, despite using glycogen at a faster rate. The glucose homeostasis defect correlates with the inability of fasted RagA(GTP/GTP) neonates to trigger autophagy and produce amino acids for de novo glucose production. Because profound hypoglycemia does not inhibit mTORC1 in RagA(GTP/GTP) neonates, Efeyan et al. (2013) considered the possibility that the Rag pathway signals glucose as well as amino acid sufficiency to mTORC1. Indeed, mTORC1 is resistant to glucose deprivation in RagA(GTP/GTP) fibroblasts, and glucose, like amino acids, controls its recruitment to the lysosomal surface, the site of mTORC1 activation. Thus, the Rag GTPases signal glucose and amino acid concentrations to mTORC1, and have an unexpectedly key role in neonates in autophagy induction and thus nutrient homeostasis and viability.
Bar-Peled, L., Chantranupong, L., Cherniack, A. D., Chen, W. W., Ottina, K. A., Grabiner, B. C., Spear, E. D., Carter, S. L., Meyerson, M., Sabatini, D. M.A tumor suppressor complex with GAP activity for the Rag GTPases that signal amino acid sufficiency to mTORC1. Science 340: 1100-1106, 2013. [PubMed: 23723238] [Full Text: https://doi.org/10.1126/science.1232044]
Deng, L., Jiang, C., Chen, L., Jin, J., Wei, J., Zhao, L., Chen, M., Pan, W., Xu, Y., Chu, H., Wang, X., Ge, X., Li, D., Liao, L., Liu, M., Li, L., Wang, P.The ubiquitination of RagA GTPase by RNF152 negatively regulates mTORC1 activation. Molec. Cell 58: 804-818, 2015. [PubMed: 25936802] [Full Text: https://doi.org/10.1016/j.molcel.2015.03.033]
Efeyan, A., Zoncu, R., Chang, S., Gumper, I., Snitkin, H., Wolfson, R. L., Kirak, O., Sabatini, D. D., Sabatini, D. M.Regulation of mTORC1 by the Rag GTPases is necessary for neonatal autophagy and survival. Nature 493: 679-683, 2013. [PubMed: 23263183] [Full Text: https://doi.org/10.1038/nature11745]
Hartz, P. A.Personal Communication. Baltimore, Md. 1/27/2015.
Jewell, J. L., Kim, Y. C., Russell, R. C., Yu, F.-X., Park, H. W., Plouffe, S. W., Tagliabracci, V. S., Guan, K.-L.Differential regulation of mTORC1 by leucine and glutamine. Science 347: 194-198, 2015. [PubMed: 25567907] [Full Text: https://doi.org/10.1126/science.1259472]
Li, Y., Kang, J., Horwitz, M. S.Interaction of an adenovirus 14.7-kilodalton protein inhibitor of tumor necrosis factor alpha cytolysis with a new member of the GTPase superfamily of signal transducers. J. Virol. 71: 1576-1582, 1997. [PubMed: 8995684] [Full Text: https://doi.org/10.1128/JVI.71.2.1576-1582.1997]
Sancak, Y., Peterson, T. R., Shaul, Y. D., Lindquist, R. A., Thoreen, C. C., Bar-Peled, L., Sabatini, D. M.The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science 320: 1496-1501, 2008. [PubMed: 18497260] [Full Text: https://doi.org/10.1126/science.1157535]
Schurmann, A., Brauers, A., Massmann, S., Becker, W., Joost, H.-G.Cloning of a novel family of mammalian GTP-binding proteins (RagA, RagB(S), RagB(1)) with remote similarity to the Ras-related GTPases. J. Biol. Chem. 270: 28982-28988, 1995. [PubMed: 7499430] [Full Text: https://doi.org/10.1074/jbc.270.48.28982]
Shen, K., Huang, R. K., Brignole, E. J., Condon, K. J., Valenstein, M. L., Chantranupong, L., Bomaliyamu, A., Choe, A., Hong, C., Yu, Z., Sabatini, D. M.Architecture of the human GATOR1 and GATOR1-Rag GTPases complexes. Nature 556: 64-69, 2018. [PubMed: 29590090] [Full Text: https://doi.org/10.1038/nature26158]
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